One of the most important problems yet to be solved in biochemistry concerns the details of how the cell can regulate its own biosynthesis. This proposal will concentrate on the molecular details of how the actual protein molecules involved in the regulatory process function. Of particular concern is the enzyme aspartate transcarbamylase which exerts allosteric control over the entire pyrimidine biosynthesis pathway. In order to determine detailed information concerning how this enzyme functions on the molecular level we are producing a series of mutant aspartate transcarbamylase molecules each having a single amino acid different from the wild-type enzyme. Production of these mutant enzymes is achieved by suppression of a nonsense codon situated in the gene which codes for the enzyme. Kinetic analysis of partial purified mutants has revealed significant alterations in the enzyme's homotropic and heterotropic properties. Correlations will be made between the biochemical properties of the mutants and the molecular level perturbations caused by the amino acid substitution in terms of the enzyme's three-dimensional structure. The data obtained from these mutants should provide details of the allosteric control of this pathway which could not be obtained in any other manner.